1. Immunology and Inflammation

Protective function and durability of mouse lymph node-resident memory CD8+ T cells

  1. Scott M Anthony
  2. Natalija Van Braeckel-Budimir
  3. Steven J Moioffer
  4. Stephanie van de Wall
  5. Qiang Shan
  6. Rahul Vijay
  7. Ramakrishna Sompallae
  8. Stacey M Hartwig
  9. Isaac J Jensen
  10. Steven M Varga
  11. Noah S Butler
  12. Hai-Hui Xue
  13. Vladimir P Badovinac  Is a corresponding author
  14. John T Harty  Is a corresponding author
  1. Department of Pathology, The University of Iowa, United States
  2. Department of Microbiology and Immunology, The University of Iowa, United States
  3. Center for Discovery and Innovation, Hackensack Meridian Health, United States
  4. Interdisciplinary Graduate Program in Immunology, The University of Iowa, United States
https://doi.org/10.7554/eLife.68662
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Cite this article
  1. Scott M Anthony
  2. Natalija Van Braeckel-Budimir
  3. Steven J Moioffer
  4. Stephanie van de Wall
  5. Qiang Shan
  6. Rahul Vijay
  7. Ramakrishna Sompallae
  8. Stacey M Hartwig
  9. Isaac J Jensen
  10. Steven M Varga
  11. Noah S Butler
  12. Hai-Hui Xue
  13. Vladimir P Badovinac
  14. John T Harty
(2021)
Protective function and durability of mouse lymph node-resident memory CD8+ T cells
eLife 10:e68662.
https://doi.org/10.7554/eLife.68662
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7 figures, 3 tables and 1 additional file

Figures

Figure 1
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CD103+Β memory CD8+Β T cells are generated in draining lymph nodesΒ (LNs) during localized but not systemic infections.

(A) C57BL/6 mice were infected IN with X31 (H3N2); mice were sacrificed 90 days post-infection; non-draining cervical lymph nodes (cLN) or lung-draining mediastinal lymph nodes (mLNs) were harvested and analyzed by flow cytometry. (B) Representative plots of % of CD69 and CD103 expression (left) in NP366 tetramer+Β IV- memory CD8+Β T cells from the cLN or mLN and cumulative data (right). nΒ =Β 3–5 mice/group. Representative of three independent experiments. Bars denote mean values, dots represent independent mice. ****p<0.0001, Students t-test. (C) Mice were seeded with 104 naive P14 cells and infected IN with either PR8-GP33 (H1N1) or Vac-GP33. 30 days post-infection, draining mLNs were isolated and CD69+Β CD103+Β P14 Trm populations were evaluated (D). Representative plots (left), cumulative data (right). Representative of two independent experiments, nΒ =Β 5 mice/group. Error bars represent meanΒ Β±Β SD. ****p<0.0001, Students t-test. (E) Mice were seeded with 104 naive P14 cells and infected IP with LCMV Armstrong. 30 days post-infection LNs (mLN and iLN) were isolated and evaluated for the frequency (F) of CD69+/CD103+Β P14 s.

Figure 1β€”source data 1

Source data for Figure 1B.

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Figure 1β€”source data 2

Source data for Figure 1D.

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Figure 2
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Lung-draining LN Trm cells mediate local protective immunity.

Mice were seeded with 104 naive or 105 3M P14 cells and IN infected with PR8-GP33 virus. At 50 days post-infection, frequency of 1M or 4M P14s were measured in the peripheral blood (A). At 100 days post-infection, mLNs were harvested and the numbers of total (B) and CD103+Β CD69+ (C) P14s were determined. Representative of three independent experiments, nΒ =Β 4–5 mice/group. Error bars represent meanΒ Β±Β SD, *p<0.05, **p<0.01, Students t-test in (A–C). LCMV challenge (D,E). NaΓ―ve, 1M, or 4M mice were infected with LCMV-Armstrong (2.0 Γ— 105 PFU/mouse i.p.); 72 hr post LCMV challenge, mLN (D) and SP (E) were harvested and individually evaluated for LCMV titers by plaque assay. FTY720 treatment impact on LCMV challenge (F). NaΓ―ve, 1M, or 4M mice were infected with LCMV-Armstrong (2.0 Γ— 105 PFU/mouse i.p.) and treated with vehicle or FTY720 daily for 72 hr. 72 hr post LCMV challenge, mLN (F) were harvested and individually evaluated for LCMV titers by plaque assay. Dotted line denotes limit of detection. One representative of 2–3 independent experiments is shown, nΒ =Β 3–5 mice/group. Error bars represent meanΒ Β±Β SEM (G) or meanΒ Β±Β SD (H). NSΒ =Β not significant, *p<0.05, one-way ANOVA.

Figure 2β€”source data 1

Source data for Figure 2A.

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Figure 2β€”source data 2

Source data for Figure 2B.

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Figure 2β€”source data 3

Source data for Figure 2C.

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Figure 2β€”source data 4

Source data for Figure 2D.

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Figure 2β€”source data 5

Source data for Figure 2E.

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Figure 2β€”source data 6

Source data for Figure 2F.

https://cdn.elifesciences.org/articles/68662/elife-68662-fig2-data6-v1.xlsx
Figure 3 with 1 supplement
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Repeated antigen stimulation extends the survival of LN Trms.

Mice were seeded with 104 naive or 105 3M P14 cells and IN infected with PR8-GP33 virus. At indicated time points, mLN (A) was harvested and total numbers of 1M (white) and 4M (black) P14 cells were evaluated. Representative of three independent experiments, nΒ =Β 4 mice/group/time point. Error bars represent meanΒ Β±Β SD. ***p<0.001, ****p<0.0001, two-way ANOVA with Sidak’s multiple comparison test. Representative plots (B) and cumulative results (C) of 1M and 4M CD69+Β CD103+Β P14 Trm cells in mLN evaluated at indicated time points. Representative of three independent experiments, nΒ =Β 4 mice/group/time point. Error bars represent meanΒ Β±Β SD. *p<0.05, **p<0.01, ****p<0.0001, two-way ANOVA with Sidak’s multiple comparison test.

Figure 3β€”source data 1

Source data for Figure 3A.

https://cdn.elifesciences.org/articles/68662/elife-68662-fig3-data1-v1.xlsx
Figure 3β€”source data 2

Source data for Figure 3C.

https://cdn.elifesciences.org/articles/68662/elife-68662-fig3-data2-v1.xlsx
Figure 3β€”figure supplement 1
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CD103+Β LN Trm cells are not present within non-draining iLN.

Mice were seeded with 104naiveΒ P14 or 105 3M P14 cells and IN infected with PR8-GP33 virus.Β At indicated time points, iLN (A) were harvested and total numbers of 1M (white) and 4M (black) P14 cells were evaluated. Representative of three independent experiments, nΒ =Β 4 mice/group/time point. Error bars represent meanΒ Β±Β SD. ***p<0.001, ****p<0.0001, two-way ANOVA with Sidak’s multiple comparison test. Representative plots of 1M and 4M CD69+Β CD103+Β P14 Trm cells in iLN (B) evaluated 30 and 150 days post-infection. Representative of three independent experiments, nΒ =Β 4 mice/group/time point.

Figure 3β€”figure supplement 1β€”source data 1

Source data for Figure 3β€”figure supplement 1A.

https://cdn.elifesciences.org/articles/68662/elife-68662-fig3-figsupp1-data1-v1.xlsx
Figure 4 with 2 supplements
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Repeated antigen stimulation increases granzyme production of LN Trms.

Mice were seeded with 104 naive or 105 3M P14 cells and IN infected with PR8-GP33 virus. AtΒ >60 days post-infection, mLNs were harvested, stimulated with cognate (GP33) peptide in the presence of BFA for 5 hr, and ICS was performed to evaluate the frequency of GrzA+Β and GrzB+Β fractions of IVβˆ’Β CD103βˆ’ or CD103+Β 1M or 4M by flow cytometry. Representative flow plots (A), cumulative frequencies (B), and total numbers per mLN (C) are shown. Representative of two independent experiments, nΒ =Β 4–5 mice/group. Error bars represent meanΒ Β±Β SD. *p<0.05, one-way ANOVA in (B), *p<0.05, Students t-test in (C).

Figure 4β€”source data 1

Source data for Figure 4B.

https://cdn.elifesciences.org/articles/68662/elife-68662-fig4-data1-v1.xlsx
Figure 4β€”source data 2

Source data for Figure 4C.

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Figure 4β€”figure supplement 1
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Repeated influenza stimulation reduces cytokine production, but does not affect degranulation capacity of LN Trm cells.

Mice were seeded with 104naiveΒ P14 or 105 3M P14 cells and IN infected with PR8-GP33 virus. At 60 days post-infection, mLN and spleen were harvested and cells analyzed for peptide-stimulated cytokine production and degranulation. (A) Gating strategy for analysis of IVβˆ’ P14s. Single-cell suspensions were stimulated with cognate peptide (GP33) in the presence of BFA for 5 hr and ICS was performed to evaluate the frequency of CD107+, IFNΞ³+, TNF+,Β and IL-2+Β fractions of IVβˆ’ CD103βˆ’ or CD103+Β 1M or 4M by flow cytometry. Representative flow plots of IFNg, TNF, and IL-2 (B), cumulative bar graphs denoting frequencies of CD107+ (C, left) and CD107+Β IFNg+Β (C, right) are displayed. Representative of two independent experiments, nΒ =Β 4–5 mice/group. NSΒ =Β not significant, *p<0.05, **p<0.01, ***p<0.001,Β ****p<0.0001; two-way ANOVA with Sidak’s multiple comparison test.

Figure 4β€”figure supplement 1β€”source data 1

Source data for Figure 4β€”figure supplement 1C.

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Figure 4β€”figure supplement 2
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Repeated antigen stimulation alters the phenotype of LN Trm cells.

Mice were seeded with 104naiveΒ P14 or 105 3M P14 cells and IN infected with PR8-GP33 virus. AtΒ >90 days post-infection, iLN (top) and mLN (bottom) were harvested and the frequency of 1M and 4M P14 cells evaluated. (A) Representative data examining CD62L expression; cumulative frequency data (middle); cumulative total number data (right). Representative of three independent experiments, nΒ =Β 4 mice/group. Error bars represent meanΒ Β±Β SD.Β ***p<0.001, ****p<0.0001 one-way ANOVA. AtΒ >75 days post-infection, mLN were harvested from 1M and 4M mice and the relative expressions of CD122 (B), Eomes (C), CD49a (D), CXCR6 (E), and CXCR3 (F) by CD103+Β (red) or CD103– (blue) P14s were evaluated. Representative histograms (top) and cumulative data (bottom) are displayed. Representative of three independent experiments, nΒ =Β 4–5 mice/group. Error bars represent meanΒ Β±Β SD. NSΒ =Β not significant, *p<0.05, ****p<0.0001, one-way ANOVA.

Figure 4β€”figure supplement 2β€”source data 1

Source data for Figure 4β€”figure supplement 2A.

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Figure 4β€”figure supplement 2β€”source data 2

Source data for Figure 4β€”figure supplement 2B.

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Figure 4β€”figure supplement 2β€”source data 3

Source data for Figure 4β€”figure supplement 2C.

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Figure 4β€”figure supplement 2β€”source data 4

Source data for Figure 4β€”figure supplement 2D.

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Figure 4β€”figure supplement 2β€”source data 5

Source data for Figure 4β€”figure supplement 2E.

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Figure 4β€”figure supplement 2β€”source data 6

Source data for Figure 4β€”figure supplement 2F.

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Figure 5
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Repeated antigen stimulation alters localization of LN Trm cells.

(A–D) Mice containing mixed congenically distinct populations of 1M (CD90.1/.1, seeded with 104 naive P14) and 4M (CD90.1/.2 seeded with 105 3M P14) P14s were injected with bolus IV administration of CD90.1-PE (red) and CD90.2-APC (white). Approximately 5 hr post-injection, organs were isolated and two-photon microscopy was performed on whole mLN (A–B) or iLN (C–D) explants ex vivo. The LN surrounding collagen capsule (pseudocolored blue) was captured with secondary harmonic generation (SHG). Representative of two independent experiments, nΒ =Β 3–4 mice/group.

Figure 5β€”source data 1

Source data for Figure 5A–D.

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Figure 6
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Residential nature of 1M and 4M LN Trm cells primed by influenza infection.

(A) 90 days after IN PR8-GP33 infection, mice bearing Thy1.1/1.1 1M P14 (green; 1M mice seeded with 104 naive P14) cells were joined by parabiotic surgery with mice bearing Thy1.1/1.2 4M P14 (purple, 4M mice seeded with 105 3M P14). Three weeks later parabionts were analyzed. (B) Abundance of 1M (green) and 4M (purple) P14 cells in iLNs of 1M (top row) and 4M (bottom row) parabiotic mice. Representative plots (left), cumulative data (right). Representative of two independent experiments, nΒ =Β 4 parabionts/experiment. Error bars represent meanΒ Β±Β SD. ****p<0.0001, t-test. (C) Abundance and distribution of 1M (green) and 4M (purple) Trm P14 cells expressed as a % of the total Trm population (CD69+/CD103+) in mLN of 1M (top row) and 4M (bottom row) parabiotic mice. Representative plots (left), cumulative data (right). Representative of two independent experiments, nΒ =Β 4 parabionts/experiment. Error bars represent meanΒ Β±Β SD. Two-way ANOVA with Sidak’s multiple comparison test. Q1(1M) vs Q1(4M) ****p<0.0001; Q2(1M) vs Q2(4M) ****p<0.0001; Q1(1M) vs Q1(4M) ****p<0.0001.

Figure 6β€”source data 1

Source data for Figure 6B.

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Figure 6β€”source data 2

Source data for Figure 6C.

https://cdn.elifesciences.org/articles/68662/elife-68662-fig6-data2-v1.xlsx
Figure 7 with 1 supplement
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Splenic 4M cells express a core Trm signature.

Mice were seeded with 104 naive P14 or 105 3M P14 cells and IN infected with PR8-GP33 virus. At 22–30 days post-infection, IV exclusion was performed and negatively enriched pooled groups of spleens (3–5 spleens/sample, nΒ =Β 3) or mLNs (15–25 mLNs/sample, nΒ =Β 2) were stained for CD8Ξ±, CD90.1, CD69, and CD103. Bulk RNAseq was performed on RNA from sort-purified spleen samples (20k IVβˆ’, CD69βˆ’/CD103βˆ’ cells/sample) or mLN Trm samples (2–5k IVβˆ’ CD69+/CD103+Β cells/sample). (A) Heatmaps of 1300 most differentially expressed genes (log2FCΒ >Β 1.5, p<0.05) between 1M and 4M LN Trms are plotted from the four respective groups of samples. The six core signature sets of genes offset to the right were derived from unbiased hierarchical clustering. (B) Volcano plot of 4061 differentially expressed genes between 1M and 4M splenic memory P14 cells (log2FCΒ >Β 1.5, p<0.05). (C) GSEA of core Trm genes defined in Table 2 from splenic 1M and 4M populations separated into respective upregulated (top) and downregulated (bottom) gene sets in regard to annotated expression in Trms. (D) Heatmap of a core set of selected Trm genes (as in C) within 1M and 4M splenic populations.

Figure 7β€”figure supplement 1
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Gating strategy of FACS for RNAseq.

Mice were seeded with 104 naive P14 or 105 3M P14 cells and IN infected with PR8-GP33 virus. At 23–30 days post-infection, mLN and spleen were pooled, harvested, negatively enriched, and sorted for bulk RNAseq. (A) Gating strategy for analysis of 1M or 4M splenic IVβˆ’ Tcirc P14s. (B) Gating strategy for analysis of 1M or 4M IVβˆ’ mLN Trm P14s.

Tables

Table 1
Samples for RNAseq, cell number, and RNA quality.
RNAseq sampleInformation
Sample info(pg/Β΅L) RNAΒ concentrationSample volume (Β΅l)Total ng,Β RNA per tube(RIN) RNA
Integrity Number asΒ determined by Agilent		RNAΒ ratio
1M Sp DN #16181710.506N/A2
1M SP DN #26781711.526N/A2
1M SP DN #310181717.306N/A2
4M Sp DN #16531711.1019.32.2
4M SP DN #26921711.7649.62.3
4M SP DN #310281717.4769.42.1
1M mLN Trms78171.3269.21.8
1M mLN Trms139233.1977.41.8
4M mLN Trms12001720.49.11.8
4M mLN Trms8181713.9068.71.8
Table 2
RNAseq heatmaps and GSEA full data.

RNAseq expression related to Figure 6.

GeneSp1M-1Sp1M-2Sp1M-3Sp4M-1Sp4M-2Sp4M-3
Neurl3βˆ’1.448372βˆ’0.7443671.4137260.0291060.5586490.191258
Ctla4βˆ’1.472581βˆ’0.893646βˆ’0.0856310.7646740.937260.749924
Fn1βˆ’0.4761βˆ’0.7844470.313751βˆ’0.47611.898997βˆ’0.4761
Arl4cβˆ’0.219927βˆ’0.637316βˆ’0.954088βˆ’0.3682041.8395740.33996
Pdcd1βˆ’0.974857βˆ’1.5210790.2718570.6475150.790750.785815
Bcl2βˆ’1.381925βˆ’0.6532171.527735βˆ’0.0806610.0199930.568075
Rgs2βˆ’1.527485βˆ’0.9837760.3442620.8180420.64780.701157
Rgs16βˆ’1.15537βˆ’1.037752βˆ’0.4692270.8871620.8360290.939158
Fcgr2bβˆ’0.959014βˆ’0.670621βˆ’1.0688590.8536310.7526441.092219
Cd244βˆ’0.4632βˆ’0.776816βˆ’1.377560.8844620.8769980.856116
Slamf7βˆ’1.139613βˆ’1.165538βˆ’0.2949170.9038170.8606080.835643
Atf3βˆ’0.779708βˆ’0.779708βˆ’0.7797080.5172440.1206881.701193
Il2raβˆ’1.322889βˆ’1.1275480.0218610.7336150.8892190.805742
Zeb2βˆ’1.316736βˆ’1.060834βˆ’0.1249860.8439570.6924070.966192
Nr4a2βˆ’1.166233βˆ’1.064171βˆ’0.3910730.701880.8612061.058391
Itga60.103598βˆ’1.3548661.208944βˆ’1.0450110.4770860.610248
Chn1βˆ’0.408248βˆ’0.408248βˆ’0.4082482.041241βˆ’0.408248βˆ’0.408248
Itga4βˆ’1.533817βˆ’0.9350030.9560910.5902660.2548590.667605
Cd44βˆ’0.687106βˆ’0.641904βˆ’1.1524410.1386321.0616341.281186
Sema6dβˆ’1.095775βˆ’1.095775βˆ’0.2772191.3041330.3151540.849482
Dusp2βˆ’1.284804βˆ’1.2482050.3633330.5169720.7117240.94098
Bcl2l1βˆ’1.599476βˆ’0.9207340.6505490.5632170.6603480.646096
Fgf13βˆ’1.184461βˆ’1.3528960.333820.5806070.8363260.786603
Cxcr3βˆ’1.558265βˆ’0.9347180.893920.3683230.6701380.560601
Fabp5βˆ’0.937223βˆ’0.580515βˆ’1.1615530.9662390.9879020.72515
Tnfsf10βˆ’0.757581βˆ’1.3349βˆ’0.5291280.7828340.7450861.093689
Rorcβˆ’0.833666βˆ’0.8336660.249621βˆ’0.8336660.7267271.52465
Cd101βˆ’1.953973βˆ’0.1224770.3286940.4243730.6217660.701617
Csf1βˆ’1.089549βˆ’0.89601βˆ’0.7350680.9050820.8727740.94277
S1pr1βˆ’1.414071βˆ’1.105490.2617920.6953590.8344680.727942
Usp33βˆ’1.470917βˆ’1.0454780.7252380.8698960.3258550.595405
Bach2βˆ’1.176816βˆ’0.990541.5231350.4171380.352504βˆ’0.125421
Aqp3βˆ’1.330326βˆ’1.2165940.8337990.3714890.7627890.578843
Coro2aβˆ’1.303957βˆ’1.114023βˆ’0.0624950.7849560.8721980.823321
Tnfsf8βˆ’1.655898βˆ’0.0802021.4780250.0135870.258145βˆ’0.013656
Dmrta1βˆ’0.845576βˆ’1.1498191.698186βˆ’0.0559750.0941920.258992
Junβˆ’1.376227βˆ’1.1624850.3131260.6818020.7650770.778706
Zfp683βˆ’1.593545βˆ’0.9021610.4625860.7343250.4911770.807619
Runx3βˆ’1.150841βˆ’1.3271610.9872230.1855120.489350.815917
Tnfrsf1bβˆ’1.135037βˆ’1.4268120.6221790.4988930.6889430.751834
Abcb1aβˆ’1.375583βˆ’1.001921βˆ’0.1094720.6917160.8660810.929179
Cd36βˆ’1.132023βˆ’1.1320230.6470121.3507240.414674βˆ’0.148364
Fosl20.574051βˆ’1.743753βˆ’0.6171940.2605880.8530120.673296
Cd38βˆ’1.726006βˆ’0.5467150.1692240.3422290.7683550.992914
Klf3βˆ’1.580977βˆ’0.9485170.6123430.6342950.6230610.659795
Cxcl9βˆ’0.779708βˆ’0.7797080.5172441.7011930.120688βˆ’0.779708
Tgfbr3βˆ’1.496104βˆ’1.0031560.2722290.6115540.8420890.773388
Dtx1βˆ’1.323911βˆ’0.5323751.651323βˆ’0.2711030.0796770.39639
Vps37bβˆ’1.327024βˆ’1.2482280.7315730.5238880.6731870.646604
Hsph1βˆ’0.89509βˆ’1.269058βˆ’0.4820190.7634310.9215050.961231
Chn2βˆ’1.469041βˆ’1.0398230.3667150.5657770.9665510.609821
Il12rb2βˆ’1.512436βˆ’0.863636βˆ’0.0441590.7171790.8419540.861097
Cd8b1βˆ’0.897526βˆ’1.057029βˆ’0.743550.6505281.072770.974808
Cd8aβˆ’1.369955βˆ’0.899035βˆ’0.2931710.6914080.9589220.91183
Sema4fβˆ’0.711384βˆ’1.581909βˆ’0.1307660.7076070.8411390.875313
Mxd1βˆ’1.636166βˆ’0.8353040.8202570.4200780.5204510.710683
Bhlhe40βˆ’1.377693βˆ’0.960477βˆ’0.1943660.7949750.8766390.860922
Klrg1βˆ’0.649531βˆ’0.92849βˆ’1.1279760.8804010.9458220.879774
Lag3βˆ’1.314079βˆ’1.2506610.476270.5621760.7368420.789452
Klre1βˆ’0.817609βˆ’0.924946βˆ’0.9914910.9008790.8931070.940061
Klrd1βˆ’0.82806βˆ’1.237129βˆ’0.6056830.7790210.844551.0473
Klrc2βˆ’1.060961βˆ’0.22419βˆ’1.1578391.3128170.3187380.811434
Klrc1βˆ’0.456844βˆ’0.628525βˆ’1.481540.7993080.8740630.893538
Dusp16βˆ’1.549192βˆ’0.590124βˆ’0.3043610.4825390.9402861.020853
Emp1βˆ’0.820283βˆ’1.020728βˆ’0.886020.7900460.9617560.975229
Fosbβˆ’1.192929βˆ’1.192929βˆ’0.1047190.8179430.6426771.029956
Nkg70.8566620.552759βˆ’1.936070.0028260.4555850.068237
Ppp1r15aβˆ’1.743276βˆ’0.6907680.6518690.5833820.597130.601662
Swap70βˆ’1.185293βˆ’1.2736270.028840.7604660.8368270.832787
Il4raβˆ’1.240991βˆ’1.2702771.0219410.408520.3806980.70011
Il21rβˆ’1.376141βˆ’1.1768480.8582310.4300210.6305420.634194
Itgalβˆ’1.41421βˆ’1.1391990.448350.5758170.778120.751121
Itgaxβˆ’1.299963βˆ’1.141223βˆ’0.0059960.7301090.8436860.873388
Bag3βˆ’1.261842βˆ’1.3025850.5799480.4716190.8192370.693622
Adam8βˆ’1.448605βˆ’0.760408βˆ’0.3550880.8022970.9089790.852826
Ifitm2βˆ’1.9315680.697426βˆ’0.2219550.3543910.5721630.529542
Ifitm1βˆ’0.116103βˆ’0.604934βˆ’1.6390240.6020240.99390.764137
Ifitm3βˆ’0.399411βˆ’1.471071βˆ’0.006047βˆ’0.1649961.5523870.489138
Ifitm10βˆ’1.619476βˆ’0.8103670.1984430.6304750.8339450.76698
Ifngr1βˆ’1.689527βˆ’0.7242360.5428520.317250.8735660.680095
Prdm1βˆ’0.763724βˆ’0.527892βˆ’1.2666071.198390.9740680.385764
Prf1βˆ’1.450717βˆ’1.0221220.1298330.7324090.8193840.791213
Gzmmβˆ’0.148129βˆ’1.7688640.896369βˆ’0.2978850.8686970.449812
Gadd45bβˆ’1.732358βˆ’0.682270.4054530.5920530.6236960.793427
Phlda1βˆ’1.336075βˆ’1.1975980.2864570.7830690.8143780.649768
Ifngβˆ’1.270643βˆ’0.812118βˆ’0.5874440.8720870.9420330.856084
A430078g2βˆ’0.768864βˆ’1.6724110.3529950.7627010.5466240.778955
Rasa3βˆ’1.298438βˆ’1.2682760.4562230.6121020.7407350.757654
Dusp4βˆ’0.013625βˆ’1.524651βˆ’0.248373βˆ’0.0136251.6047240.19555
Lplβˆ’0.751462βˆ’0.751462βˆ’0.751462βˆ’0.0584560.5961171.716724
Klf2βˆ’0.890686βˆ’1.058738βˆ’0.6228390.3040691.262581.005613
Inpp4bβˆ’1.411831βˆ’1.1229560.9179490.5487020.4084870.65965
Dnajb1βˆ’1.351654βˆ’1.1870330.503080.4560150.9562110.623381
Junbβˆ’1.248952βˆ’1.3241030.753570.5085820.6790450.631859
Cdh1βˆ’1.503072βˆ’0.8545781.2390260.4980730.178620.441931
Itgb1βˆ’1.070625βˆ’1.246026βˆ’0.2450610.6698110.9083230.98358
Fut11βˆ’1.566705βˆ’0.9073430.2644370.6404650.7106430.858503
Gzmcβˆ’1.129587βˆ’1.1295871.3937450.7015020.0819630.081963
Gzmbβˆ’1.019828βˆ’0.852547βˆ’0.8603090.8606050.9368480.935231
Tmem123βˆ’0.486668βˆ’0.568492βˆ’1.3832660.2892731.3328770.816276
Icam1βˆ’1.395117βˆ’1.1776560.6226490.5928310.7357180.621575
S1pr5βˆ’1.401126βˆ’1.0706170.1072560.687380.8265630.850543
Gm10080βˆ’0.187622βˆ’0.228053βˆ’1.7977390.5849550.7341870.894272
Smad3βˆ’1.296473βˆ’1.2540740.4365540.4947560.8176830.801554
Anxa2βˆ’1.161681βˆ’1.045596βˆ’0.4440540.7777620.9357550.937813
Cx3cr1βˆ’1.267574βˆ’1.170718βˆ’0.0230680.7782250.8460350.837099
Ccr8βˆ’1.030745βˆ’1.030745βˆ’0.6493170.8594990.9020930.949215
Crr9βˆ’1.163685βˆ’1.1636850.5181730.0593331.3692190.380645
Cxcr6βˆ’1.008609βˆ’0.713182βˆ’0.9986560.855040.9865420.878867
Ccr1βˆ’0.959366βˆ’0.959366βˆ’0.044992βˆ’0.4601371.1489551.274906
Ccr2βˆ’1.244353βˆ’0.427864βˆ’0.8351070.6475980.4702741.389451
Ccr5βˆ’1.267395βˆ’1.1977810.0848130.6002190.8304190.949725
Adam19βˆ’1.693324βˆ’0.3535171.3074940.4079040.361716βˆ’0.030273
Havcr2βˆ’0.267637βˆ’1.661471βˆ’0.4792170.7470140.8198720.841438
Itgae0.265873βˆ’0.131851βˆ’1.8906340.2878541.0309810.437777
Traf4βˆ’1.26168βˆ’1.3170190.6640030.6564930.7005340.557669
Ccl50.7178050.069708βˆ’1.9657560.1554630.6964470.326333
Ccl9βˆ’0.096406βˆ’1.204224βˆ’1.1769340.6856581.0956860.69622
Ccl30.3217410.441288βˆ’2.0338210.3472050.5573510.366236
Ccl4βˆ’0.900786βˆ’1.071795βˆ’0.7437440.7708110.9760860.969428
Wfikkn2βˆ’1.521007βˆ’0.4642331.525421βˆ’0.0379380.1519380.345819
Tbx21βˆ’1.188999βˆ’1.375880.4753450.6103450.7590990.720091
Arl5cβˆ’1.071685βˆ’1.4812440.6221130.5211950.7559160.653704
Stat3βˆ’1.33085βˆ’1.2448270.5684210.5758720.7221030.709281
Icam2βˆ’1.427743βˆ’1.0742860.3079950.5011990.9659030.726932
Cmahβˆ’1.619268βˆ’0.8200811.0124380.4813390.4439190.501653
Fam65bβˆ’1.435265βˆ’1.0876510.2783840.7565490.736230.751753
Irf4βˆ’0.942848βˆ’1.402093βˆ’0.1664210.7376520.8741450.899565
Ly86βˆ’1.176696βˆ’1.1766960.2714780.7022550.0930941.286564
Nfil3βˆ’1.352986βˆ’1.2106740.4426960.6649490.7807730.675242
Cdc14bβˆ’1.660352βˆ’0.625526βˆ’0.0189910.6824790.6244630.997927
Naip3βˆ’0.645497βˆ’0.645497βˆ’0.645497βˆ’0.6454971.2909941.290994
Elovl7βˆ’0.792302βˆ’0.420558βˆ’1.3505781.1072280.9343820.521828
Gzmaβˆ’0.686737βˆ’0.814558βˆ’1.1971820.8487370.9410360.908703
Gzmkβˆ’1.406303βˆ’0.870165βˆ’0.2784020.7648090.911270.878791
Itga1βˆ’1.285811βˆ’1.102208βˆ’0.1132170.8349490.7035020.962785
Rhobβˆ’0.230141βˆ’1.8690040.1305540.3631980.712530.892863
Id2βˆ’1.046589βˆ’0.909419βˆ’0.7604470.7602240.9149861.041244
Adam40.587047βˆ’1.9859930.480645βˆ’0.0352460.5870470.3665
Fosβˆ’0.879435βˆ’1.290058βˆ’0.4740260.8821880.816450.94488
Il7rβˆ’0.170366βˆ’0.087313βˆ’1.8259590.4237860.640091.019763
Ly6c1βˆ’0.753065βˆ’1.398044βˆ’0.2146370.1747511.0678361.123158
Ly6c2βˆ’1.276681βˆ’1.2094690.159690.5481360.8961530.882171
Il2rbβˆ’1.273008βˆ’1.302080.7056620.5058920.6653740.698162
Bin2βˆ’1.441078βˆ’1.1226640.5463090.6004360.7095770.70742
Nr4a1βˆ’1.212606βˆ’1.3122521.0272330.5543660.4620630.481197
Itga5βˆ’0.995646βˆ’0.9956461.538635βˆ’0.3969880.6811750.16847
Litafβˆ’1.67839βˆ’0.6921260.1531890.8174330.8273320.572561
Klhl6βˆ’1.527286βˆ’0.9083520.1379480.5615230.8883020.847865
Bcl6βˆ’1.042258βˆ’1.315998βˆ’0.1400060.8510541.0252310.621977
Tigitβˆ’0.636695βˆ’0.492403βˆ’1.4529250.801080.9728090.808135
Sidt1βˆ’1.416356βˆ’0.9179451.3523630.3209310.3085180.352489
Ccr6βˆ’0.408248βˆ’0.408248βˆ’0.408248βˆ’0.4082482.041241βˆ’0.408248
Hspa1aβˆ’1.489338βˆ’1.057890.5465750.592070.6008620.807722
Tnfβˆ’1.932303βˆ’0.2261970.372220.5082530.6663420.611686
Tnfsf9βˆ’0.721049βˆ’1.122053βˆ’0.8693070.8717380.995940.84473
Qpctβˆ’0.9723320.746934βˆ’0.972332βˆ’0.7738320.9191291.052433
Slc3a2βˆ’1.412175βˆ’1.036270.0688120.6417120.9199150.818006
Dtx4βˆ’1.8958210.0310870.9854320.2063820.6235260.049393
Dusp5βˆ’1.204462βˆ’1.231643βˆ’0.0095550.6557360.907920.882003
Aff30.3341580.9341361.302955βˆ’0.695519βˆ’1.131655βˆ’0.744075
Icos1.4711370.8413930.145304βˆ’0.792242βˆ’1.047218βˆ’0.618376
Ikzf2βˆ’0.1656711.7049780.658251βˆ’0.793835βˆ’0.609887βˆ’0.793835
Cxcr40.8691150.9193330.906884βˆ’0.886976βˆ’1.182769βˆ’0.625587
Cd550.6944670.4440261.397141βˆ’0.459842βˆ’1.074231βˆ’1.001562
Rgs10.282992βˆ’0.0288891.3206630.669655βˆ’1.444527βˆ’0.799896
Sell0.3430050.5773991.57097βˆ’0.879322βˆ’0.741027βˆ’0.871025
Xcl11.1580690.7267650.823087βˆ’0.933921βˆ’0.948108βˆ’0.825893
Slamf60.8781150.6480261.158166βˆ’0.969274βˆ’0.688009βˆ’1.027024
Dapl10.958760.9531710.822988βˆ’0.894936βˆ’0.945047βˆ’0.894936
Mal0.278549βˆ’1.1663880.89838βˆ’0.0247421.180589βˆ’1.166388
Pmepa11.0009431.530105βˆ’0.429028βˆ’0.700673βˆ’0.700673βˆ’0.700673
Fabp4βˆ’0.4082482.041241βˆ’0.408248βˆ’0.408248βˆ’0.408248βˆ’0.408248
Skil1.478111.07101βˆ’0.815616βˆ’0.553424βˆ’0.577221βˆ’0.602859
Il20.8430971.080960.786459βˆ’0.913213βˆ’0.735725βˆ’1.061578
Foxo11.3980041.047696βˆ’0.261312βˆ’1.167157βˆ’0.508615βˆ’0.508615
Il6ra0.1924770.1232631.813008βˆ’0.551343βˆ’0.551343βˆ’1.026062
Npr1βˆ’0.4082482.041241βˆ’0.408248βˆ’0.408248βˆ’0.408248βˆ’0.408248
Lef1βˆ’0.577413βˆ’0.1225822.008562βˆ’0.513026βˆ’0.534901βˆ’0.26064
Lpar3βˆ’0.600795βˆ’0.6007951.779753βˆ’0.600795βˆ’0.6007950.623426
Ifi44βˆ’1.5221881.3705390.77727βˆ’0.414406βˆ’0.162093βˆ’0.049122
Nr4a31.658887βˆ’1.4347770.298688βˆ’0.213395βˆ’0.096008βˆ’0.213395
Tlr11.087014βˆ’1.7597010.693949βˆ’0.332077βˆ’0.046720.357535
Cxcl100.819849βˆ’1.5536171.2170930.268993βˆ’0.179362βˆ’0.572955
Cd27βˆ’0.8127020.7360121.686726βˆ’0.76644βˆ’0.35287βˆ’0.490726
Cd91.1398530.9630040.584072βˆ’0.83021βˆ’0.83021βˆ’1.02651
Cd690.6246430.930421.125309βˆ’0.853084βˆ’0.712926βˆ’1.114363
Isg201.3934780.9346870.197602βˆ’0.930029βˆ’0.737091βˆ’0.858648
Rgs100.966341.0376460.595869βˆ’0.585759βˆ’0.644411βˆ’1.369686
Themisβˆ’0.6592091.957135βˆ’0.786652βˆ’0.207908βˆ’0.035484βˆ’0.267882
Fyn1.0665841.3465470.054683βˆ’0.726098βˆ’0.837588βˆ’0.904128
Egr21.036188βˆ’1.6368371.073379βˆ’0.057847βˆ’0.260015βˆ’0.154868
Timp3βˆ’0.4082482.041241βˆ’0.408248βˆ’0.408248βˆ’0.408248βˆ’0.408248
Dusp6βˆ’0.2059910.0851391.646718βˆ’1.49606βˆ’0.014903βˆ’0.014903
Crtam1.0792031.145370.408387βˆ’1.043006βˆ’0.745838βˆ’0.844116
Cxcr5βˆ’1.4752450.9949791.2045650.116051βˆ’0.50998βˆ’0.330371
Eomesβˆ’0.0051120.3622011.517082βˆ’1.5599440.048375βˆ’0.362601
Tcf7βˆ’0.5629370.0666071.970539βˆ’0.522208βˆ’0.289205βˆ’0.662796
Ccr7βˆ’0.893954βˆ’0.1633251.969748βˆ’0.395389βˆ’0.21246βˆ’0.30462
Cd861.3808651.183911βˆ’0.546108βˆ’0.52407βˆ’0.777066βˆ’0.717531
Btla0.9717060.8747180.860138βˆ’0.76399βˆ’1.171717βˆ’0.770855
Sik10.1951640.1951640.1951641.1168090.195164βˆ’1.897467
Epcam1.0638440.4984880.93633βˆ’1.165034βˆ’1.165034βˆ’0.168594
Egr1βˆ’0.013398βˆ’0.6993011.896141βˆ’0.6993010.151141βˆ’0.635282
Ifit31.1146240.96780.615644βˆ’0.896586βˆ’0.812924βˆ’0.988558
Key resources table
Reagent type (species) or resourceDesignationSource or referenceIdentifiersAdditional
information
Strain, strain background (Mus musculus)C57BL6/JJackson LaboratoryStock No: 000664
(RRID:IMSR_JAX:000664)
Strain, strain background (Mus musculus)B6.PL(84NS)/CyJackson LaboratoryStock No: 000983
(RRID:IMSR_JAX:000406)		C57BL6/J
Thy1.1
Strain, strain background (Mus musculus)B6.Cg-Tcratm1MomTg(TcrLCMV)327Sdz
(P14)		Jackson LaboratoryStock No: 37394-JAX
(RRID:IMSR_TAC:4138)
Strain, strain background (Mus musculus)Thy1.1/1.1- B6.Cg-Tcratm1Mom Tg(TcrLCMV)327SdzThis paperThy1.1/1.1 P14Can be acquired through lab contact or breeding of above commercially available strains
Strain, strain background (Mus musculus)Thy1.1/1.2- B6.Cg-Tcratm1Mom Tg(TcrLCMV)327SdzThis paperThy1.1/1.2 P14Can be acquired through lab contact or breeding of above commercially available strains
Strain, strain background (Influenza A virus)Recombinant influenza A/PR/8/34 expressing (H1N1) GP33-41Laidlaw et al. Cooperativity Between CD8+ T Cells, Non- Neutralizing Antibodies, and Alveolar Macrophages Is Important for Heterosubtypic Influenza Virus Immunity. Plos Pathog. 9(3) e1003207 (2013).PR8-GP33Can be acquired through lab contact.
Strain, strain background (Influenza A virus)Recombinant influenza A/X-31 (H3N2) expressing GP33-41Laidlaw et al. Cooperativity Between CD8+ T Cells, Non- Neutralizing Antibodies, and Alveolar Macrophages Is Important for Heterosubtypic Influenza Virus Immunity. Plos Pathog. 9(3) e1003207 (2013).X31-GP33Can be acquired through lab contact.
Peptide, recombinant proteinGP33-44AnaSpecCatalog #:
AS-61296
AntibodyCD11a
(rat monoclonal)		BiolegendM17/4
(AB_312776)		FACs
(1:100)
AntibodyIFNΞ³
(rat monoclonal)		eBioscienceXMG1.2
(AB_465410)		FACs
(1:100)
AntibodyCD8a
(rat monoclonal)		eBioscience53–6.7
(AB_1853141)		FACs
(1:100)
AntibodyThy1.1
(mouse monoclonal)		eBioscienceOX-7
(AB_2201314)		FACs
(1:100)
AntibodyThy1.2
(rat monoclonal)		eBioscience30-H12
(AB_1853152)		FACs
(1:100)
AntibodyCD45.2
(mouse monoclonal)		eBioscience104
(AB_469724)		FACs
(1:100)
AntibodyCD103
(hamster monoclonal)		Biolegend2E7
(AB_469040)		FACs
(1:100)
AntibodyCD69
(hamster monoclonal)		BiolegendH1.2F3
(AB_1853105)		FACs
(1:100)
AntibodyCD44
(rat monoclonal)		Biolegend1M7
(AB_223593)		FACs
(1:100)
AntibodyCD62L
(rat monoclonal)		BiolegendMEL-14
(AB_1853103)		FACs
(1:100)
AntibodyKLRG1
(mouse monoclonal)		eBioscience2F1
(AB_540279)		FACs
(1:100)
AntibodyCX3CR1
(mouse monoclonal)		eBioscienceSA011F11
(AB_2565701)		FACs
(1:100)
AntibodyCXCR3
(Armenian hamster monoclonal)		eBioscienceCXCR3-173
(AB_1210593)		FACs
(1:100)
AntibodyEomesodermin
(rat monoclonal)		eBioscienceDan11mag
(AB_11042577)		FACs
(1:100)
AntibodyTNF
(rat monoclonal)		eBioscienceMP6-XT22
(AB_465416)		FACs
(1:100)
AntibodyIL-2
(rat monoclonal)		BiolegendJES6-5H4
(AB_315298)		FACs
(1:100)
AntibodyGranzyme A
(mMouse monoclonal)		Biolegend3G8.5
(AB_2565308)		FACs
(1:100)
AntibodyGranzyme B
(rat monoclonal)		Biolegend12F9B65
(AB_2564373)		FACs
(1:100)
AntibodyBrdU
(mouse monoclonal)		BiolegendBu20a
(AB_1595472)		FACs
(1:100)
Commercial assay or kitFoxp3/Transcription Factor Staining Buffer SetInvitrogen00-5523-00
Software, algorithmGraphPad PrismGraphPad Prism 8Version 8.4.2 (464)
(RRID:SCR_002798)

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  1. Scott M Anthony
  2. Natalija Van Braeckel-Budimir
  3. Steven J Moioffer
  4. Stephanie van de Wall
  5. Qiang Shan
  6. Rahul Vijay
  7. Ramakrishna Sompallae
  8. Stacey M Hartwig
  9. Isaac J Jensen
  10. Steven M Varga
  11. Noah S Butler
  12. Hai-Hui Xue
  13. Vladimir P Badovinac
  14. John T Harty
(2021)
Protective function and durability of mouse lymph node-resident memory CD8+ T cells
eLife 10:e68662.
https://doi.org/10.7554/eLife.68662